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The genetic diversity of sperm and spermatangia stored by mated Japanese flying squid (Todarodes pacificus) was examined to better understand how this ommastrephid squid mates and stores sperm. Analyses were conducted using spermatangia implanted in the lips of immature squid collected in Tsugaru Strait, northern Japan, in July–August 2019, spermatangia implanted in the lips of mature squid collected in the East China Sea in April 2019, and sperm stored in the seminal receptacles of squid collected off Tosashimizu, Kochi Prefecture, Japan, in February 2019. DNA was extracted, microsatellite loci were amplified using five PCR primers, and allele peaks were analysed. Analysis of the spermatangia confirmed that females in the field mate with multiple males, suggesting that males transfer from 1 to at least 14 spermatangia in each mating. Analysis of the seminal receptacles showed that sperm from multiple males are mixed within individual sperm chambers. Spermatangia were also found implanted in the lips of males, providing evidence of same-sex sexual behaviour. To date, studies of cephalopod mating systems have focused on coastal species, and the results of this study add to our growing knowledge of more oceanic species.

The life history of copepods during nauplius stages has rarely been studied, because of difficulties in identification. This study describes the development of an identification method that uses integrated microscopic analysis and real-time PCR based method to distinguish and estimate the abundance of pelagic calanoida nauplii, Neocalanus plumchrus, and to determine their phenology and ontogenetic vertical migration (OVM) in the Oyashio region. The overall approach of identification included: classify field samples of calanoida nauplii into six developmental stages by microscopic analysis, determining the Cq values from each representative nauplii by real-time PCR using specie specific probe and primer pair for N. plumchrus, calculating the |ΔCq| values from the standard curve, distinguish N. plumchrus nauplii by the criterion of |ΔCq| ⩽ 2, and calculate the abundance of N. plumchrus nauplii using results obtained from microscopic analysis and real-time PCR. Validation of the method showed that our real-time PCR based approach could identify N. plumchrus nauplii in field samples with as low as 9% of underestimation, the effect of which is negligibly low on the investigation. The results showed that early nauplius stages (first and second nauplius stages: N1 and N2, respectively) were observed during the period from October 2009 to May 2010. Spawning females were also observed during the same period, and these results confirm that the spawning period of N. plumchrus is at least 8 months long. N3 was the most dominant stage in October 2009, January, and March 2010, the distribution depth of N3 was between 500 m and 2000 m in October 2009 and their vertical distribution range was extended until April 2010. Late stages (N4–N6) were rarely observed during the period from October 2009 to March 2010, and they occurred abundantly from the surface layer (0–20 m) in April 2010. The dominance of N3 from October to March and the sudden occurrence of later stages in April indicate arrested development of N3 in the mesopelagic layer.